Lindemans, J.

A periconceptional energy-rich dietary pattern is associated with early fetal growth: The Generation R study (Article)

2012-12-04T00:00:00Z

Objective: To identify periconceptional maternal dietary patterns associated with crown-rump length (CRL), estimated fetal weight (EFW) and birthweight. Design: Population-based prospective birth cohort study. Setting: Rotterdam, the Netherlands. Participants: For this study, 847 pregnant Dutch women were eligible. Women were included between 2001 and 2005. Methods: Information on nutritional intake was collected by a semiquantitative food frequency questionnaire. For extracting dietary patterns, principal component factor analysis was used. Fetal growth was assessed using ultrasound measurements. Information on birth outcomes was retrieved from medical records. Multivariate regression analyses were used. Main outcome measures: Crown-to-rump length, estimated fetal weight in second and third trimester and birthweight. Results: An 'energy-rich dietary pattern' was identified, characterised by high intakes of bread, margarine and nuts. A significant association was shown between a high adherence to this dietary pattern (difference, mm: 2.15, 95% confidence interval 0.79-3.50) and CRL (linear trend analyses P = 0.015). No association was revealed between increasing adherence to this dietary pattern and EFW in second or third trimester, or birthweight. Conclusion: This study suggests that increasing adherence to an energy-rich dietary pattern is associated with increased CRL in the first trimester.

Peptide fingerprinting of folate-responsive proteins in human B lymphoblasts and orofacial clefting (Article)

2012-07-01T00:00:00Z

Background Maternal periconceptional use of folic acid contributes to the prevention of neural crest-related congenital malformations including orofacial clefts. The underlying biological pathways affected by folic acid, however, are still not clarified. In an explorative study, we identify folate-responsive proteins and pathways by advanced proteomic techniques and their possible role in orofacial development in young children. Materials and methods At 15months of age, we obtained B lymphoblasts from 10 children with and 10 children without an orofacial cleft. Folate-responsive protein expression was determined in folate-free B-lymphoblast cultures, supplemented with 5-methyltetrahydrofolate to reach the target concentration 30nM. Folate-associated differences of peptide and protein expressions were assessed by analysing samples before and after folate addition. Samples were trypsin digested and measured by nano-liquid chromatography coupled online to a LTQ-Orbitrap mass spectrometer. Significantly differentiating peptides were determined using a McNemar's test, and correlations with proteins and existing pathways were visualized using Ingenuity Pathway Analysis. Results We found 39 folate-responsive peptides that were assigned to 30 proteins. Those proteins consisted of histones, ribosomal and heat shock proteins (HSP), and proteins involved in antioxidant reactions, cytoskeleton, glycolysis, energy production, protein processing, signal transduction and translation. Conclusions Histones, ribosomal and HSP were mainly found in the case group, and we confirm that almost 60% of these proteins were also found in a subset of the samples in our previous study using microarray on folate-responsive gene expression. The proteins were compared with known biological pathways and matched with recent relevant literature.

Comparable levels of folate-induced aneusomy in B-lymphoblasts from oral-cleft patients and controls (Article)

2012-01-24T00:00:00Z

Background: Peri-conceptional use of folic acid contributes to protection against congenital malformations, such as neural tube defects and cleft lip with or without cleft palate (CL/P). Previous studies showed that low folate levels cause DNA damage, leading to chromosomal instability and aneusomy. This study seeks to confirm this finding and investigates whether the in vitro sensitivity towards aneusomy of chromosome 17 and 21 in the folate-deficient state differs between CL/P patients and controls. Methods: Epstein-Barr virus-immortalized B-lymphoblasts derived from 15 CL/P children and 15 controls, were cultured in medium with high and low concentrations - approximately 40. nM and 5. nM - of 5-methyltetrahydrofolate, respectively. Fluorescence in situ hybridization was used to detect specific fluorescence signals for chromosomes 17 and 21. Results: A significant increase in aneusomy of chromosomes 17 (2.3% vs 7.6%; p≤ 0.001) and 21 (2.5% vs 7.0%; p≤ 0.001) was observed after 10 days of culturing in low folate. These results were comparable in cell lines from patients and controls. Interestingly, for chromosome 17 the folate deficiency mainly resulted in an increase of monosomy (6%, p≤ 0.001), while for chromosome 21 the increase of trisomy was larger (4.9%, p≤ 0.001). Conclusions: These data suggest that folate deficiency is a significant risk factor in the development of aneusomy and may affect the distribution of chromosomes during cell division. The comparable aneusomy frequencies in CL/P and in controls suggest that other folate-related processes are involved in the pathogenesis of CL/P, and additional investigations are needed to identify the causal mechanisms.

Mass spectrometry analyses of κ and λ fractions result in increased number of complementarity-determining region identifications (Article)

2012-01-01T00:00:00Z

Sera from lung cancer patients contain antibodies against tumor-associated antigens. Specific amino acid sequences of the complementarity-determining regions (CDRs) in the antigen-binding fragment (Fab) of these antibodies have potential as lung cancer biomarkers. Detection and identification of CDRs by mass spectrometry can significantly be improved by reduction of the complexity of the immunoglobulin molecule. Our aim was to molecular dissect IgG into κ and λ fragments to reduce the complexity and thereby identify substantially more CDRs than by just total Fab isolation. We purified Fab, Fab-κ, Fab-λ, κ and λ light chains from serum from 10 stage I lung adenocarcinoma patients and 10 matched controls from the current and former smokers. After purification, the immunoglobulin fragments were enzymatically digested and measured by high-resolution mass spectrometry. Finally, we compared the number of CDRs identified in these immunoglobulin fragments with that in the Fab fragments. Twice as many CDRs were identified when Fab-κ, Fab-λ, κ and λ (3330) were combined than in the Fab fraction (1663) alone. The number of CDRs and κ:λ ratio was statistically similar in both cases and controls. Molecular dissection of IgG identifies significantly more CDRs, which increases the likelihood of finding lung cancer-related CDR sequences.

C-reactive protein levels, blood pressure and the risks of gestational hypertensive complications: The Generation R Study (Article)

2011-12-01T00:00:00Z

OBJECTIVES: Aim of this study was to investigate the associations of C-reactive protein levels, as marker of low-grade inflammation, with blood pressure development during pregnancy and the risks of gestational hypertensive complications. We also explored the role of maternal BMI in these associations. METHODS: High-sensitivity C-reactive protein levels were measured in early pregnancy (median 13.2 weeks, 95% range 9.6-17.6) in 5816 mothers participating in a population-based prospective cohort study in the Netherlands. Blood pressure measurements were performed in each trimester. Information about pregnancy-induced hypertension and preeclampsia was retrieved from hospital charts of the women. RESULTS: Longitudinal analyses showed that C-reactive protein levels were not associated with SBP and DBP patterns throughout pregnancy. Trimester-specific multivariate linear regression models showed that as compared to low C-reactive protein levels (<5.0 mg/l), elevated levels (≥20.0 mg/l) were associated with maternal SBP and DBP. Elevated C-reactive protein levels in early pregnancy were associated with the risks of pregnancy-induced hypertension [odds ratio (OR) 2.78, 95% confidence interval (CI) 1.66-4.66]. After adjustment for maternal BMI, all associations attenuated. CONCLUSION: Our results suggest that first-trimester C-reactive protein levels are associated with SBP and DBP levels throughout pregnancy and with gestational hypertensive complications, but these associations are largely explained by maternal BMI.

Assessment of intracellular methotrexate and methotrexate-polyglutamate metabolite concentrations in erythrocytes by ultrafast matrix-assisted laser desorption/ionization triple quadrupole tandem mass spectrometry (Article)

2011-10-30T00:00:00Z

A new ultrafast quantitative and high-throughput mass spectrometric method using matrix-assisted laser desorption/ionization triple quadrupole tandem mass spectrometry has been developed and validated for determination of intracellular erythrocyte concentrations of the antifolate drug methotrexate (MTX) and its polyglutamate metabolites. The method consists of a solid-phase extraction of MTX and MTX-polyglutamate metabolites from deproteinized erythrocyte lysates spiked with aminopterin as internal standard. The newly developed method was validated according to the most recent FDA guidelines on linearity, recovery, within-run and between-run accuracy and precision and stability of the analytes. The low limit of quantification (LLOQ) was 10 nmol/L for all analytes while the limit of detection (LOD) determined at a signal-to-noise (S/N) ratio = 3:1 in drug- free erythrocyte lysate was on average 0.3 nmol/L. After validation, the new method was used in the measurement of intracellular erythrocyte concentrations of MTX and MTX-polyglutamate metabolites (MTXPG2 to MTXPG7) in packed human erythrocyte samples collected from patients with rheumatoid arthritis receiving low-dose oral methotrexate therapy. Mean (SD) intracellular erythrocyte concentrations observed in patient samples were 12.8 (12.6), 12.4 (9.4), 44.4 (30.0), 33.6 (35.9) and 9.4 (8.2) nmol/L for MTX to MTXPG5, respectively, in 106erythrocytes. The highest observed glutamylation degree of MTX was MTXPG5, the very long chain MTX-polyglutamate metabolites MTXPG6 and MTXPG7 were not detected in the packed erythrocyte pellets collected from rheumatoid arthritis patients.

Tailored preconceptional dietary and lifestyle counselling in a tertiary outpatient clinic in the Netherlands (Article)

2011-09-01T00:00:00Z

Background Adverse reproductive performance has been linked to unhealthy dietary intake and lifestyles. Our objectives were to investigate the prevalence of unhealthy dietary intake and lifestyles before conception and to evaluate whether tailored preconception counselling modifies these behaviours. Methods Between October 2007 and April 2009, 419 couples received tailored preconception dietary and lifestyle counselling at the outpatient clinic of Obstetrics and Gynaecology of the Erasmus University Medical Center Rotterdam, the Netherlands. A subgroup (n = 110 couples) was counselled twice with a fixed time interval of 3 months. Self-administered questionnaires were used for tailored dietary and lifestyle counselling. A cumulative score based on six Dutch dietary guidelines was displayed in the personal Preconception Dietary Risk score (PDR score). In a similar manner, the Rotterdam Reproduction Risk score (R3 score) was calculated from lifestyle factors (women: 13 items, men: 10 items). Univariate and paired tests were used. Results Most couples (93.8) were subfertile. At the second counselling, the percentage consuming the recommended intake of fruit had increased from 65 to 80 in women and from 49 to 68 in men and the percentage of women getting the recommended intake of fish increased from 39 to 52. As a consequence, the median PDR score was decreased [women: 2.6 (95 CI 2.4-2.9) to 2.4 (95 CI 2.1-2.6), men: 2.5 (95 CI 2.3-2.7) to 2.2 (95 CI 1.9-2.4), both P < 0.05]. The median R3 scores were also lower [women: 4.7 (95 CI 4.3-5.0) to 3.1 (95 CI 2.8-3.4), men: 3.0 (95 CI 2.8-3.3) to 2.0 (95 CI 1.7-2.3), both P < 0.01] due to less alcohol use (-14.6), more physical exercise and folic acid use in women, and less alcohol use in men (-19.4) (all P < 0.01). The R3 scores in women and men were decreased in all ethnicity, educational level, neighbourhood and BMI categories. However, low educated women appeared to show a larger reduction than better educated women and men with a normal BMI to show a larger decrease than overweight men. The reduction in the PDR score of women was similar in both ethnic groups. More than 85 women and men found the counselling useful and around 70 would recommend it to others. Conclusions Tailored preconception counselling about unhealthy dietary and lifestyle behaviours of subfertile couples in an outpatient tertiary clinic is feasible and seems to decrease the prevalence of harmful behaviours in the short term. These Results with subfertile couples are promising and illustrate their opportunities to contribute to reproductive performance and pregnancy outcome.

Major dietary patterns and blood pressure patterns during pregnancy: the Generation R Study (Article)

2011-08-18T00:00:00Z

Objective: We sought to evaluate associations between dietary patterns and systolic blood pressure (SBP) and diastolic blood pressure during pregnancy. Study design: This was a prospective study of 3187 pregnant women. Participants completed a food-frequency questionnaire in early pregnancy. The Mediterranean dietary pattern, comprising high intake of vegetables, vegetable oils, pasta, fish, and legumes, and the Traditional dietary pattern, comprising high intake of meat and potatoes, were identified using factor analysis. Results: A higher SBP was observed among mothers with high Traditional pattern adherence. Low adherence to the Mediterranean pattern was also associated with higher SBP but only in early and mid pregnancy. A higher diastolic blood pressure throughout pregnancy was observed in mothers with high adherence to the Traditional pattern and low adherence to the Mediterranean pattern. These effect estimates were most pronounced in mid pregnancy. Conclusion: Low adherence to a Mediterranean and high adherence to a Traditional dietary pattern is associated with a higher blood pressure in pregnancy.

Analytical investigations of toxic p-phenylenediamine (PPD) levels in clinical urine samples with special focus on MALDI-MS/MS (Article)

2011-08-10T00:00:00Z

Para-phenylenediamine (PPD) is a common chromophoric ingredient in oxidative hair-dyes. In some African countries like Sudan, Egypt and Morocco but also in India this chemical is used alone or in combination with colouring extracts like Henna for dyeing of the hair or the skin. Excessive dermal exposure to PPD mainly leads to the N-mono- and N,N′-diacetylated products (MAPPD, DAPPD) by N-acetyltransferase 1 and 2 (NAT1 and 2) catalyzed reactions. Metabolites and PPD are mainly excreted via renal clearance. Despite a low risk of intoxication when used in due form, there are numerous cases of acute intoxication in those countries every year. At the ENT Hospital - Khartoum (Sudan) alone more than 300 cases are reported every year (~10% fatal), mostly caused by either an accidental or intended (suicidal) high systemic exposure to pure PPD. Intoxication leads to a severe clinical syndrome including laryngeal edema, rhabdomyolysis and subsequent renal failure, neurotoxicity and acute toxic hepatitis. To date, there is no defined clinical treatment or antidote available and treatment is largely supportive. Herein, we show the development of a quick on-site identification assay to facilitate differential diagnosis in the clinic and, more importantly, the implementation of an advanced analytical platform for future in-depth investigations of PPD intoxication and metabolism is described. The current work shows a sensitive (~25 μM) wet chemistry assay, a validated MALDI-MS/MS and HPLC-UV assay for the determination of PPD and its metabolites in human urine. We show the feasibility of the methods for measuring PPD over a range of 50-1000 μM. The validation criteria included linearity, lower limit of quantification (LLOQ), accuracy and precision, recovery and stability. Finally, PPD concentrations were determined in clinical urine samples of cases of acute intoxication and the applied technique was expanded to identify MAPPD and DAPPD in the identical samples.

C-reactive protein levels in early pregnancy, fetal growth patterns, and the risk for neonatal complications: The Generation R Study (Article)

2011-08-01T00:00:00Z

Objective: We sought to examine the associations of maternal C-reactive protein (CRP) levels with fetal growth and the risks of neonatal complications. Study Design: CRP levels were measured in early pregnancy in 6016 women. Main outcome measures were fetal growth in each trimester and neonatal complications. Results: As compared to the reference group (CRP levels <5 mg/L), elevated maternal CRP levels (<25 mg/L) were associated with lower estimated fetal weight in third trimester and lower weight at birth (differences: -29 g, 95% confidence interval [CI], -58 to 0 and -128 g, 95% CI, -195 to -60, respectively). Elevated maternal CRP levels were also associated with an increased risk of a small size for gestational age in the offspring (adjusted odds ratio, 2.94; 95% CI, 1.615.36). Conclusion: Maternal CRP levels in early pregnancy are associated with fetal growth restriction and increased risks of neonatal complications.

The CYP2C19*2 genotype predicts tamoxifen treatment outcome in advanced breast cancer patients (Article)

2011-08-01T00:00:00Z

Aims: Tamoxifen is metabolized by cytochrome P450s, with an important role for CYP2D6. Recently, we demonstrated in 80 patients that CYP2C19*2 is associated with increased survival in breast cancer patients using tamoxifen. Here, we aimed to confirm this in a large group of 499 patients. Materials & methods: A total of 499 estrogen receptor-positive primary breast tumor specimens of advanced disease patients treated with first-line tamoxifen were genotyped for CYP2C19*2 and 17 variant alleles, with primary end point time-to-treatment failure (TTF). Effects of CYP2C19, independent of treatment, were analyzed in 243 primary systematic untreated patients. Results: CYP2C19*2 hetero-and homozygote patients combined showed significantly longer TTFs (hazard ratio [HR]: 0.72; 95% CI: 0.57-0.90; p = 0.004). In multivariate analysis, including CYP2D6*4 status, CYP2C19*2 remained independently associated with TTF (HR: 0.73; 95% CI: 0.58-0.91; p = 0.007). In untreated patients, the CYP2C19*17 allele was significantly associated with a longer disease-free interval (HR: 0.66; 95%CI: 0.46-0.95; p = 0.025). Conclusion: CYP2C19 genotyping is potentially important for tamoxifen therapy for advanced disease and for breast cancer prognosis.

Effects of rosuvastatin on progression of stenosis in adult patients with congenital aortic stenosis (PROCAS Trial) (Article)

2011-07-15T00:00:00Z

Recent trials have failed to show that statin therapy halts the progression of calcific aortic stenosis (AS). We hypothesized that statin therapy in younger patients with congenital AS would be more beneficial, because the valve is less calcified. In the present double-blind, placebo-controlled trial, 63 patients with congenital AS (age 18 to 45 years) were randomly assigned to receive either 10 mg of rosuvastatin daily (n = 30) or matched placebo (n = 33). The primary end point was the progression of peak aortic valve velocity. The secondary end points were temporal changes in the left ventricular mass, ascending aortic diameter, and N-terminal pro-brain natriuretic peptide (NT-proBNP). The median follow-up was 2.4 years (interquartile range 1.9 to 3.0). The mean increase in peak velocity was 0.05 ± 0.21 m/s annually in the rosuvastatin group and 0.09 ± 0.24 m/s annually in the placebo group (p = 0.435). The annualized change in the ascending aorta diameter (0.4 ± 1.7 mm with rosuvastatin vs 0.5 ± 1.6 mm with placebo; p = 0.826) and left ventricular mass (1.1 ± 15.8 g with rosuvastatin vs -3.7 ± 30.9 g with placebo; p = 0.476) were not significantly different between the 2 groups. Within the statin group, the NT-proBNP level was 50 pg/ml (range 19 to 98) at baseline and 21 pg/ml (interquartile range 12 to 65) at follow-up (p = 0.638). NT-proBNP increased from 40 pg/ml (interquartile range 20 to 92) to 56 pg/ml (range 26 to 130) within the placebo group (p = 0.008). In conclusion, lipid-lowering therapy with rosuvastatin 10 mg did not reduce the progression of congenital AS in asymptomatic young adult patients. Interestingly, statins halted the increase in NT-proBNP, suggesting a potential positive effect of statins on cardiac function in young patients with congenital AS.

Ultrafast selective quantification of methotrexate in human plasma by high-throughput MALDI-isotope dilution mass spectrometry (Article)

2011-06-01T00:00:00Z

Background: A new analytical MS method using isotope dilution combined with MALDI-triple quadrupole MS/MS has been developed and validated for the determination of methotrexate and 7-hydroxymethotrexate in plasma. Methotrexate, methotrexate-d3, 7-hydroxymethotrexate and 7-hydroxymethotrexate-d3 were monitored by selected reaction monitoring using the transitions m/z 455.2→308.2, 458.2→311.2, 471.2→324.2 and 474.2→327.2 for methotrexate, methotrexate-d3, 7-hydroxymethotrexate and 7-hydroxymethotrexate- d3, respectively. Results: The LLOQ was 1 nmol/l for methotrexate and 7-hydroxymethotrexate while the limit of detection was 0.3 nmol/l for both analytes. The new developed method was cross-validated by a fluorescence polarization immunoassay and tested for its clinical feasibility by measuring plasma samples from patients suffering from acute lymphoblastic leukemia. Plasma methotrexate concentrations ranged between 66.0 and 954 nmol/l and observed 7-hydroxymethotrexate/methotrexate ratios ranged between 0.1 and 32.4, respectively. Conclusion: The new method showed comparable analytical performances as the fluorescence polarization immunoassay, but analyte specificity and sensitivity of the newly developed method were significantly better.

Preconception folic acid use modulates estradiol and follicular responses to ovarian stimulation (Article)

2011-02-01T00:00:00Z

Background: Folate is a methyl donor. Availability of folate affects DNA methylation profiles and thereby gene expression profiles. We investigated the effects of low-dose folic acid use (0.4 mg/d) on the ovarian response to mild and conventional ovarian stimulation in women. Methods: In a randomized trial among subfertile women, 24 and 26 subjects received conventional and mild ovarian stimulation, respectively. Blood samples were taken during the early follicular phase of the cycle prior to treatment and on the day of human chorionic gonadotropin administration for determination of serum total homocysteine, anti-Müllerian hormone (AMH), estradiol, and folate. Folic acid use was validated by questionnaire and serum folate levels. Preovulatory follicles were visualized, counted, and diameters recorded using transvaginal ultrasound. The relation between folic acid use and ovarian response was assessed using linear regression analysis. Results: Folic acid use modified the ovarian response to ovarian stimulation treatment. The estradiol response was higher in nonfolic acid users receiving conventional treatment [βinteraction=0.52 (0.07- 0.97); P = 0.03], and this effect was independent of serum AMH levels and the preovulatory follicle count. In the conventional treatment, themeanfolliclenumberwasalso greater in nonusers compared with the users group (14.1 vs. 8.9, P = 0.03). Conclusion: Low-dose folic acid use attenuates follicular and endocrine responses to conventional stimulation, independent of AMH and follicle count. The nature of this observation suggests that the effect of folic acid is most prominent during early follicle development, affecting immature follicles. Deleterious effects of folate deficiency, like DNA hypomethylation and oxidative stress, can help to explain our observations.

A derangement of the maternal lipid profile is associated with an elevated risk of congenital heart disease in the offspring (Article)

2010-12-27T00:00:00Z

Background and aims: Maternal hyperglycaemia and hyperhomocysteinaemia are risk factors for congenital heart disease (CHD). These metabolic derangements and deranged lipid levels are associated with adult cardiovascular disease. We examined whether maternal lipid levels are associated with the risk of CHD offspring. Methods and Results: From 2003 onwards, a case-control study was conducted. Participants were mothers of children with (n = 261) and without (n = 325) CHD. At around 16 months after the index-pregnancy, maternal lipid levels were determined. Maternal characteristics and lipid levels were compared by Student's t-test. In a multivariable logistic regression model, risk estimates were calculated for associations between CHD and lipid levels. Adjustments were made for maternal age, diabetes, ethnicity, body mass index (BMI), parity, periconception folic acid use and total homocysteine levels. Outcome measures are presented in (geometric) means (p5-p95) and odds ratios (ORs) with 95% confidence intervals (CIs). Case mothers showed higher cholesterol (4.9 vs. 4.7 mmol l-1, P < 0.05), low-density lipoprotein (LDL)-cholesterol (3.2 vs. 3.0 mmol l-1, P < 0.05), apolipoprotein B (84.0 vs. 80.0 mg dl-1, P < 0.01) and homocysteine (10.8 vs. 10.2 μmol l-1, P < 0.05) than controls. LDL-cholesterol above 3.3 mmol l-1(OR 1.6 (95%CI, 1.1-2.3)) and apolipoprotein B above 85.0 mg dl-1were associated with an almost twofold increased CHD risk (OR 1.8 (95%CI, 1.2-2.6)). This was supported by elevated CHD risks per unit standard deviation increase in cholesterol (OR 1.2 (95% CI 1.03-1.5)), LDL-cholesterol (OR 1.3 (95%CI, 1.1-1.6) and apolipoprotein B (OR 1.3 (95% CI 1.1-1.6)). Apolipoprotein B was most strongly associated with CHD risk. Conclusion: A mildly deranged maternal lipid profile is associated with an increased risk of CHD offspring.

The preconception nutritional status of women undergoing fertility treatment: Use of a one-year post-delivery assessment (Article)

2010-12-01T00:00:00Z

Background&Aims: A poor maternal nutritional status in the preconception period is associated with adverse pregnancy outcomes. A valid standardized assessment period after pregnancy reflecting the preconception nutritional status is missing. Therefore, this study aimed to validate the assessment period at around 1 year after delivery in women undergoing fertility treatment. Methods: In a prospective study including 30 women with a fertility problem, we compared nutrient intakes from a food frequency questionnaire and biomarkers related to the homocysteine pathway in blood, at two assessment periods, i.e., preconceptionally and 1 year after delivery. We used a linear mixed model and adjusted for possible confounders, such as body mass index and folic acid supplement use. Results: The energy-adjusted nutrient intakes were not significantly different between the two assessment periods, except for higher retinol, alcohol and vitamin B2 and lower carbohydrate intakes at around 1 year after delivery. The intraclass correlation coefficients of the nutrients ranged from 0.3 to 0.7. After adjustment, none of the biomarkers was significantly different between the two assessment periods. The intraclass correlation coefficients of the biomarkers were all ≥0.5. Conclusions: An assessment at around 1 year after delivery seems to adequately reflect the preconception nutritional status of women with a fertility problem, however larger confirmatory studies are required.

The preconception nutritional status of women undergoing fertility treatment: Use of a one-year post-delivery assessment (Article)

2010-12-01T00:00:00Z

A new ultrafast and high-throughput mass spectrometric approach for the therapeutic drug monitoring of the multi-targeted anti-folate pemetrexed in plasma from lung cancer patients (Article)

2010-12-01T00:00:00Z

An analytical assay has been developed and validated for ultrafast and high-throughput mass spectrometric determination of pemetrexed concentrations in plasma using matrix assisted laser desorption/ionization-triple quadrupole-tandem mass spectrometry. Patient plasma samples spiked with the internal standard methotrexate were measured by multiple reaction monitoring. The detection limit was 0.4 fmol/μL, lower limit of quantification was 0.9 fmol/μL, and upper limit of quantification was 60 fmol/μL, respectively. Overall observed pemetrexed concentrations in patient samples ranged between 8.7 (1.4) and 142.7 (20.3)∈pmol/μL (SD). The newly developed mass spectrometric assay is applicable for (routine) therapeutic drug monitoring of pemetrexed concentrations in plasma from non-small cell lung cancer patients.

Amino-terminal pro-brain natriuretic peptide (NT-proBNP) is a biomarker of cardiac filling pressures in pre-eclampsia (Article)

2010-11-01T00:00:00Z

Objective: To evaluate if amino-terminal pro-brain natriuretic peptide (NT-proBNP) plasma levels reflect intracardiac filling pressures in pre-eclamptic patients. Study design: In a cross-sectional study we investigated 22 untreated critically ill pre-eclamptic women between 22 and 34 weeks gestation. All patients underwent intra-arterial blood pressure and central hemodynamic measurements and NT-proBNP was determined in stored plasma. Baseline characteristics, plasma NT-proBNP concentrations and relevant laboratory variables were investigated for correlations with hemodynamic values using Spearman's rank correlation test. Results: No significant correlations were demonstrated between NT-proBNP concentrations and variables associated with the severity of the pre-eclampsia. We found significant positive correlations between NT-proBNP and diastolic pulmonary pressure (r = 0.59; p = 0.005) and pulmonary capillary wedge pressure (PCWP) (r = 0.51; p = 0.015). Multiple linear regression analysis showed that the association between NT-proBNP and PCWP was not affected by creatinine level. Conclusion: NT-proBNP is a biomarker of left ventricular cardiac filling pressures in untreated pre-eclamptic patients.

Retinol status of newborn infants is associated with congenital diaphragmatic hernia (Article)

2010-10-01T00:00:00Z

OBJECTIVE: Genetic analyses in humans suggest a role for retinoid-related genes in the pathogenesis of congenital diaphragmatic hernia (CDH). The goal of this study was to investigate the vitamin A status of mothers and their newborns in association with CDH. METHODS: We conducted a hospital-based, case-control study with 22 case and 34 control mothers and their newborns. In maternal and cord blood samples, retinol and retinol-binding protein (RBP) levels were measured with high-performance liquid chromatography and an enzyme-linked immunosorbent assay, respectively. Univariate and multivariate logistic regression analyses were performed to determine crude and adjusted risk estimates. RESULTS: Case newborns had significantly lower levels of retinol (0.60 vs 0.76 μmol/L; P=.003) and RBP (5.42 vs 7.11 mg/L; P=.02) than did control newborns. The multivariate logistic regression analysis showed lower levels of retinol and RBP in association with CDH risk; the odds ratio for retinol levels of <15th percentile (<0.61 μmol/L) was 11.11 (95% confidence interval: 2.54-48.66; P=.001), and that for RBP levels of <15th percentile (<4.54 mg/L) was 4.00 (95% confidence interval: 1.00 -15.99; P=.05). Retinol and RBP levels were not different between case and control mothers. CONCLUSIONS: CDH is strongly associated with low retinol and RBP levels in newborns, independent of maternal retinol status. This is an important finding supporting the idea that human CDH is linked with abnormal retinoid homeostasis.

Arterial stiffness as determinant of increased amino terminal pro-B-type natriuretic peptide levels in individuals with and without cardiovascular disease - The Rotterdam Study (Article)

2010-10-01T00:00:00Z

Objective: Large artery stiffening has adverse effects on cardiac structure and function and, therefore, may be associated with elevated circulating levels of amino-terminal pro-B-type natriuretic peptide (NT-proBNP). Methods: In a large community-dwelling older population (n = 6211, mean age 69.2 years), serum NT-proBNP, brachial pulse pressure (PP) and carotid-femoral pulse wave velocity (cfPWV) were measured. Results: In individuals without cardiovascular disease (CVD), median NT-proBNP was 6.7 pmol/l in men (n = 2073) and 10.1 pmol/l in women (n = 3085) (P < 0.001). In these individuals, indices of arterial stiffness correlated with NT-proBNP with beta-coefficients for brachial PP and cfPWV of 0.315 and 0.255 in men and 0.233 and 0.232 in women (all P < 0.001). After multivariable adjustment (age, weight, height, mean arterial pressure, heart rate, smoking, diabetes, estimated glomerular filtration rate, total and high-density lipoprotein-cholesterol and use of lipid-lowering and antihypertensive medication), these associations remained significant for brachial PP and cfPWV in men and for brachial PP in women. In multivariable-adjusted models, brachial PP explained 20.3% and cfPWV 10.7% of the variation of NT-proBNP in men and, respectively, 10.8 and 9.4% in women. In patients with prevalent CVD, indices of arterial stiffness and NT-proBNP were unrelated in multivariable-adjusted models. Conclusion: Our findings show that arterial stiffness is independently associated with elevated NT-proBNP levels in individuals without prevalent CVD. The association between vascular stiffness and NT-proBNP is stronger in men than in women and absent in individuals with prevalent CVD.

Prognosis of Vascular Surgery Patients Using a Quantitative Assessment of Troponin T Release: Is the Crystal Ball still Clear? (Article)

2010-09-29T00:00:00Z

Background: Cardiac troponin T (cTnT) assays with increased sensitivity might increase the number of positive tests. Using the area under the curve (AUC) with serial sampling of cTnT an exact quantification of the myocardial damage size can be made. We compared the prognosis of vascular surgery patients with integrated cTnT-AUC values to continuous and standard 12-lead electrocardiography (ECG) changes. Methods: 513 Patients were monitored. cTnT sampling was performed on postoperative days 1, 3, 7, 30 and/or at discharge or whenever clinically indicated. If cTnT release occurred, daily measurements of cTnT were performed, until baseline was achieved. CTnT-AUC was quantified and divided in tertiles. All-cause mortality and cardiovascular events (cardiac death and myocardial infarction) were noted during follow-up. Results: 81/513 (16%) Patients had cTnT release. After adjustment for gender, cardiac risk factors, and site and type of surgery, those in the highest cTnT-AUC tertile were associated with a significantly worse cardiovascular outcome and long-term mortality (HR 20.2; 95% CI 10.2-40.0 and HR 4.0; 95% CI 2.0-7.8 respectively). Receiver operator analysis showed that the best cut-off value for cTnT-AUC was <0.01 days*ng m for predicting long-term cardiovascular events and all-cause mortality. Conclusion: In vascular surgery patients quantitative assessment of cTnT strongly predicts long-term outcome.

Risk factors and outcome of new-onset cardiac arrhythmias in vascular surgery patients (Article)

2010-06-01T00:00:00Z

Background: The pathophysiology of new-onset cardiac arrhythmias is complex and may bring about severe cardiovascular complications. The relevance of perioperative arrhythmias during vascular surgery has not been investigated. The aim of this study was to assess risk factors and prognosis of new-onset arrhythmias during vascular surgery. Methods: A total of 513 vascular surgery patients, without a history of arrhythmias, were included. Cardiac risk factors, inflammatory status, and left ventricular function (LVF; N-terminal pro-B-type natriuretic peptide and echocardiography) were assessed. Continuous electrocardiography (ECG) recordings for 72 hours were used to identify ischemia and new-onset arrhythmias: atrial fibrillation, sustained ventricular tachycardia, supraventricular tachycardia, and ventricular fibrillation. Logistic regression analysis was applied to identify preoperative risk factors for arrhythmias. Cox regression analysis assessed the impact of arrhythmias on cardiovascular event-free survival during 1.7 years. Results: New-onset arrhythmias occurred in 55 (11%) of 513 patients: atrial fibrillation, ventricular tachycardia, supraventricular tachycardia, and ventricular fibrillation occurred in 4%, 7%, 1%, and 0.2%, respectively. Continuous ECG showed myocardial ischemia and arrhythmias in 17 (3%) of 513 patients. Arrhythmia was preceded by ischemia in 10 of 55 cases. Increased age and reduced LVF were risk factors for the development of arrhythmias. Multivariate analysis showed that perioperative arrhythmias were associated with long-term cardiovascular events, irrespective of the presence of perioperative ischemia (hazard ratio 2.2, 95% CI 1.3-3.8, P = .004). Conclusion: New-onset perioperative arrhythmias are common after vascular surgery. The elderly and patients with reduced LVF show arrhythmias. Perioperative continuous ECG monitoring helps to identify this high-risk group at increased risk of cardiovascular events and death.

Evaluation of the new body fluid mode on the Sysmex XE-5000 for counting leukocytes and erythrocytes in cerebrospinal fluid and other body fluids (Article)

2010-05-01T00:00:00Z

Background: We evaluated the body fluid (BF) mode on the new Sysmex XE-5000 analyzer. Methods: Red (RBC) and white blood cell (WBC) (differential) counts of BFs (139 patient samples and 87 normal samples) were measured and compared to the Fuchs-Rosenthal chamber and stained cytospin slides. Results: Extended cell counting using the BF mode was noted to have an improved WBC detection limit (CV20%) of 10Ã106/L. Excellent agreement with the manual method was observed for most BFs [mean bias +2 to 6Ã106/L for cerebrospinal fluid (CSF) and -1 to 12Ã106/L for other fluids]. In CSF, the BF-mode counted more WBC (polymorphic nuclear cells) compared with the manual method (mean bias +5 to 6Ã106/L), especially in samples with low cell counts (<20Ã 106/L). Carry over was negligible (mostly <0.17%) and linearity was excellent (mean bias <5%). The reference ranges for CSF (n=87) were RBC 0Ã106/L, WBC and mononuclear <7Ã106/L, and polymorph nucleated cells <3Ã106/L. Conclusions: The BF mode on the Sysmex XE-5000 offers rapid and accurate RBC and WBC (differential) counts in clinically relevant concentration ranges in CSF and other fluids. In addition, the exclusion of high fluorescent cells, such as mesothelial cells and macrophages from WBC counting may reduce the number of manual analyses in pleural fluids and ascites.

Amino-terminal Pro-B-type natriuretic peptide improves cardiovascular and cerebrovascular risk prediction in the population: The rotterdam study (Article)

2010-03-01T00:00:00Z

Increased circulating amino-terminal pro-B-type natriuretic (NT-proBNP) levels are a marker of cardiac dysfunction but also associate with coronary heart disease and stroke. We aimed to investigate whether increased circulating NT-proBNP levels have additive prognostic value for first cardiovascular and cerebrovascular events beyond classic risk factors. In a community-based cohort of 5063 participants free of cardiovascular disease, aged ≥55 years, circulating NT-proBNP levels and cardiovascular risk factors were measured. Participants were followed for the occurrence of first major fatal or nonfatal cardiovascular event. A total of 420 participants developed a first cardiovascular event (108 fatal). After adjustment for classic risk factors, the hazard ratio for cardiovascular events was 2.32 (95% CI: 1.55 to 2.70) in men and 3.08 (95% CI: 1.91 to 3.74) in women for participants with NT-proBNP in the upper compared with the lowest tertile. Corresponding hazard ratios for coronary heart disease, heart failure, and ischemic stroke were 2.01 (95% CI: 1.14 to 2.59), 2.90 (95% CI: 1.33 to 4.34), and 2.06 (95% CI: 0.91 to 3.18) for men and 2.95 (95% CI: 1.30 to 4.55), 5.93 (95% CI: 2.04 to 11.2), and 2.07 (95% CI: 1.00 to 2.97) for women. Incorporation of NT-proBNP in the classic risk model significantly improved the C-statistic both in men and women and resulted in a net reclassification improvement of 9.2% (95% CI: 3.5% to 14.9%; P=0.001) in men and 13.3% (95% CI: 5.9% to 20.8%; P<0.001) in women. We conclude that, in an asymptomatic older population, NT-proBNP improves risk prediction not only of heart failure but also of cardiovascular disease in general beyond classic risk factors, resulting in a substantial reclassification of participants to a lower or higher risk category.

Periconceptional maternal folic acid use of 400 μg per day is related to increased methylation of the IGF2 gene in the very young child (Article)

2009-11-16T00:00:00Z

Background: Countries worldwide recommend women planning pregnancy to use daily 400 mg of synthetic folic acid in the periconceptional period to prevent birth defects in children. The underlying mechanisms of this preventive effect are not clear, however, epigenetic modulation of growth processes by folic acid is hypothesized. Here, we investigated whether periconceptional maternal folic acid use and markers of global DNA methylation potential (S-adenosylmethionine and S-adenosylhomocysteine blood levels) in mothers and children affect methylation of the insulin-like growth factor 2 gene differentially methylation region (IGF2 DMR) in the child. Moreover, we tested whether the methylation of the IGF2 DMR was independently associated with birth weight. Methodology/Principal Findings: IGF2 DMR methylation in 120 children aged 17 months (SD 0.3) of whom 86 mothers had used and 34 had not used folic acid periconceptionally were studied. Methylation was measured of 5 CpG dinucleotides covering the DMR using a mass spectrometry-based method. Children of mother who used folic acid had a 4.5% higher methylation of the IGF2 DMR than children who were not exposed to folic acid (49.5% vs. 47.4%; p = 0.014). IGF2 DMR methylation of the children also was associated with the S-adenosylmethionine blood level of the mother but not of the child (+1.7% methylation per SD S-adenosylmethionine; p = 0.037). Finally, we observed an inverse independent association between IGF2 DMR methylation and birth weight (-1.7% methylation per SD birthweight; p = 0.034). Conclusions: Periconceptional folic acid use is associated with epigenetic changes in IGF2 in the child that may affect intrauterine programming of growth and development with consequences for health and disease throughout life. These results indicate plasticity of IGF2 methylation by periconceptional folic acid use.

Associations between dietary patterns and semen quality in men undergoing IVF/ICSI treatment (Article)

2009-06-01T00:00:00Z

Background: This study investigates whether dietary patterns, substantiated by biomarkers, are associated with semen quality. Methods: In 161 men of subfertile couples undergoing in vitro fertilization treatment in a tertiary referral clinic in Rotterdam, the Netherlands, we assessed nutrient intakes and performed principal component factor analysis to identify dietary patterns. Total homocysteine (tHcy), folate, vitamin B12 and B6 were measured in blood and seminal plasma. Semen quality was assessed by sperm volume, concentration, motility, morphology and DNA fragmentation index (DFI). Linear regression models analyzed associations between dietary patterns, biomarkers and sperm parameters, adjusted for age, body mass index (BMI), smoking, vitamins and varicocele. Results: The 'Health Conscious' dietary pattern shows high intakes of fruits, vegetables, fish and whole grains. The 'Traditional Dutch' dietary pattern is characterized by high intakes of meat, potatoes and whole grains and low intakes of beverages and sweets. The 'Health Conscious' diet was inversely correlated with tHcy in blood (β = -0.07, P = 0.02) and seminal plasma (β = -1.34, P = 0.02) and positively with vitamin B6 in blood (β = 0.217, P = 0.01). An inverse association was demonstrated between the 'Health Conscious' diet and DFI (β = -2.81, P = 0.05). The 'Traditional Dutch' diet was positively correlated with red blood cell folate (β = 0.06, P = 0.04) and sperm concentration (β = 13.25, P = 0.01). Conclusions: The 'Health Conscious' and 'Traditional Dutch' dietary pattern seem to be associated with semen quality in men of subfertile couples.

IVF outcomes are associated with biomarkers of the homocysteine pathway in monofollicular fluid (Article)

2009-05-01T00:00:00Z

BACKGROUNDMaternal hyperhomocysteinemia is detrimental for reproduction, but the effects on embryo quality are unknown. The aim of this study was to investigate whether biomarkers of the homocysteine pathway are associated with in vitro fertilization (IVF) outcome.METHODSIn a prospective study, we investigated biomarkers of the homocysteine pathway for associations with embryo quality and biochemical pregnancy in women undergoing IVF or intracytoplasmic sperm injection treatment (n = 181). In the treatment cycle, blood and monofollicular fluid samples were collected for determination of folate, cobalamin and total homocysteine (tHcy) concentrations.RESULTSOf all the women in the study, 67 used folic acid supplements. In blood, a significant correlation was established between high cobalamin and better embryo quality [standardized adjusted regression coefficient: -0.17, 95 confidence interval (CI): -0.30, -0.01]. In monofollicular fluid of non-supplemented women, high cobalamin correlated with better embryo quality (estimate: -0.87; 95 CI: -1.68, -0.06), whereas high tHcy resulted in poor embryo quality (estimate: 1.01; 95 CI: 0.08, 1.95). However, in monofollicular fluid of supplemented women, high tHcy correlated with better embryo quality (estimate: -0.58; 95 CI: -1.12, -0.04). In the total group, a 2-fold increase of monofollicular fluid folate corresponded with a 3.3 times higher chance (95 CI: 1.09, 9.71) of achieving pregnancy.CONCLUSIONSAn optimal homocysteine pathway in follicular fluid is associated with a better embryo quality and chance of pregnancy.

Polymorphisms in folate-related genes and risk of pediatric acute lymphoblastic leukemia (Article)

2009-03-05T00:00:00Z

Polymorphisms in folate pathway genes may influence the susceptibility to acute lymphoblastic leukemia (ALL). DNA was isolated from 245 pediatric ALL patients (cases) and from 500 blood bank donors (controls). Polymorphisms in methylene-tetrahydrofolate reductase (MTHFR677C>T,1298A>C), methionine synthase (MTR 2756A>G), methionine synthase reductase (MTRR 66A>G), methylenetetrahydrofolate dehydrogenase (MTHFD1 1958G>A), nicotinamide N-methyltransferase (NNMT IVS-151C>T), serine hydroxymethyl transferase (SHMT1 1420C>T), thymidylate synthase (TS 2R3R), and the reduced folate carrier (RFC1 80G>A) were detected. In ALL patients, an increased occurrence was observed of the RFC1 80AA variant (odds ratio[OR] = 2.1; 95% confidence interval[CI] = 1.3-3.2; P =.002) and the RFC1 80A allele (OR = 1.5; 95% CI, 1.1-2.1; P =.02). Likewise, the NNMT IVS-151TT genotype showed a 2.2-fold increased ALL risk (OR = 2.2; 95% CI, 1.1-4.6; P =.04). A 1.4-fold reduction in ALL risk was observed for (heterozygous or homozygous) carriers of the TS 2R allele and the MTHFR 677T allele (OR = 0.7; 95% CI, 0.5-1.0; P <.05). Furthermore, interactions between NNMT and MTHFR 677C>T and RFC were observed. NNMT IVS-151CC/MTHFR 677CT + TT patients exhibited a 2-fold reduction in ALL risk whereas RFC1 80AA/NNMT IVS-151CT + TT subjects had a 4.2-fold increase in ALL risk (P =.001). For the first time, we associate the RFC1 80G>A and NNMT IVS-151C> T variants to an increased ALL susceptibility.(Blood. 2009;113:2284-2289)

The preconception Mediterranean dietary pattern in couples undergoing in vitro fertilization/intracytoplasmic sperm injection treatment increases the chance of pregnancy (Article)

2009-01-01T00:00:00Z

Objective: To investigate associations between preconception dietary patterns and IVF/intracytoplasmic sperm injection (ICSI) outcomes validated by biomarkers of the homocysteine pathway. Design: Observational prospective study. Setting: A tertiary referral fertility clinic at the Erasmus University Medical Centre, Rotterdam, The Netherlands. Patient(s): One hundred sixty-one couples undergoing IVF/ICSI treatment. Intervention(s): No interventions other than the Dutch governmental recommendation of folic acid. Main Outcome Measure(s): Dietary patterns, blood and follicular fluid concentrations of folate, vitamin B12, vitamin B6, homocysteine, and fertilization rate, embryo quality, and pregnancy. Result(s): In women, two dietary patterns were identified. The "health conscious-low processed" dietary pattern (variation explained 12.1%) was characterized by high intakes of fruits, vegetables, fish, and whole grains and low intakes of snacks, meats, and mayonnaise, and positively correlated with red blood cell folate (β = 0.07). The "Mediterranean" dietary pattern (variation explained 9.1%), that is, high intakes of vegetable oils, vegetables, fish, and legumes and low intakes of snacks, was positively correlated with red blood cell folate (β = 0.13), and vitamin B6 in blood (β = 0.09) and follicular fluid (β = 0.18). High adherence by the couple to the "Mediterranean" diet increased the probability of pregnancy, odds ratio 1.4 (95% confidence interval 1.0-1.9). Conclusion(s): A preconception "Mediterranean" diet by couples undergoing IVF/ICSI treatment contributes to the success of achieving pregnancy. © 2010 American Society for Reproductive Medicine.

Methionine loading does not enhance the predictive value of homocysteine serum testing for all-cause mortality or major adverse cardiac events (Article)

2009-01-01T00:00:00Z

Background: Hyperhomocysteinaemia is independently associated with atherosclerotic disease. Methionine loading could improve the predictive value of hyperhomocysteinaemia by detecting mild disturbances in enzyme activity. The aims of this study were to determine the beneficial effect of methionine loading on the predictive value of homocysteine testing for long-term mortality and major adverse cardiac events (MACE). Methods: In an observational study, 1122 patients with suspected or known vascular disease, underwent homocysteine testing, which was measured fasting and again 6 h after methionine loading. Hyperhomocysteinaemia was defined as a fasting level ≥15 μmol/L and post-methionine loading level ≥45 μmol/L or an increase of ≥30 μmol/L above fasting levels. Primary end-points were death and MACE. Multivariate Cox regression analysis was used, adjusting for all cardiac risk factors. Results: During follow up (mean 8.9 ± 3.4 years), 98 patients died (8.7%), 86 had a MACE (7.7%), 579 patients had normal tests, 134 patients had only fasting hyperhomocysteinaemia, 226 only post-methionine hyperhomocysteinaemia and 183 patients had both. In multivariate analysis, overall survival and MACE-free survival were significantly worse for those with fasting hyperhomocysteinaemia, with hazard ratios of 1.86 (95% confidence interval (CI) 1.20-2.87) and 2.24 (95%CI 1.41-3.53), respectively. The addition of hyperhomocysteinaemia after methionine loading did not significantly increase the risk of death or MACE, with hazard ratios of 0.97 (95%CI 0.52-1.81) and 0.89 (95%CI 0.47-1.69), respectively. Conclusion: The presence of post-methionine hyperhomocysteinaemia did not significantly alter risk of death or MACE in patients with normal or increased fasting homocysteine levels, respectively. In conclusion, methionine loading does not improve the predictive value of homocysteine testing with regard to long-term mortality or MACE.

Homocysteine metabolism in the pre-ovulatory follicle during ovarian stimulation (Article)

2008-11-01T00:00:00Z

BACKGROUND: Ovarian stimulation gives rise to supraphysiological estradiol levels, which may affect oocyte quality. This study aims to investigate whether ovarian stimulation deranges the homocysteine pathway thereby affecting the pre-ovulatory follicle. METHODS: Blood samples were collected on cycle day 2 and the day of hCG administration in 181 women undergoing ovarian stimulation for IVF. In each subject, the diameter of the two leading follicles was measured and the corresponding follicular fluids were collected. In blood and follicular fluid samples, total homocysteine (tHcy), folate, cobalamin and pyridoxal'5-phosphate (PLP) were determined. According to the blood folate levels, women were classified as either folic acid supplemented (n = 113) or non-supplemented (n = 32). RESULTS: Ovarian stimulation resulted in a significant decrease in blood tHcy and cobalamin levels (both P ≤ 0.001). The blood concentrations of tHcy, folate, cobalamin and PLP were significantly correlated with the corresponding follicular fluid concentrations (all P ≤ 0.001). Follicular fluid tHcy concentrations were inversely correlated with follicular diameter (P ≤ 0.05). In folic acid supplemented women, follicular fluid folate was inversely correlated with follicular diameter (P ≤ 0.05). CONCLUSIONS: Ovarian stimulation deranges blood and follicular fluid biomarkers of the homocysteine pathway. High ovarian follicular fluid tHcy and folate levels may have detrimental effects on follicular development.

Genome-wide pathway analysis of folate-responsive genes to unravel the pathogenesis of orofacial clefting in man (Article)

2008-09-01T00:00:00Z

BACKGROUND: A cleft of the lip with or without the palate (CLP) is a frequent congenital malformation with a heterogeneous etiology, for which folic acid supplementation has a protective effect. To gain more insight into the molecular pathways affected by natural folate, we examined gene expression profiles of cultured B-lymphoblasts from CLP patients before and after the addition of 5-methyltetrahydrofolate (5-mTHF) to the cultures. METHODS: Immortalized B-lymphoblasts from five children with CLP were cultured in folate-deficient medium for 5 days. 5-mTHF was added to a concentration of 30 nM. Gene expression patterns were then evaluated before and after supplementation using Human Genome U133 Plus 2.0 arrays. Data analysis was performed with Omniviz and the GEPAS analysis suite. Differential genes were categorized into biological pathways with Ingenuity Pathway systems. Differential expression was validated by quantitative RT-PCR. RESULTS: Using supervised clustering, with a false discovery rate <1%, we identified 144 and 409 significantly up-regulated and down-regulated probesets, respectively, after 5-mTHF addition. The regulated genes were involved in a variety of biological pathways, including one carbon pool and cell cycle regulation, biosynthesis of amino acids and DNA/RNA nucleotides, protein processing, apoptosis, and DNA repair. CONCLUSIONS: The large variety of the identified folate responsive pathways fits with the modifying role of folate via the methylation pathway. From the present data we may conclude that folate deficiency deranges normal cell development, which might contribute to the development of CLP. The role of these folate responsive genes in CLP development is intriguing and needs further investigation.

N-terminal pro-brain natriuretic peptide testing in the emergency department: Beneficial effects on hospitalization, costs, and outcome (Article)

2008-07-01T00:00:00Z

Background: N-terminal pro-brain natriuretic peptide (NT-proBNP) is an established biomarker for heart failure. Assessment of this biomarker in patients with acute dyspnea presenting to the emergency department (ED) may aid diagnostic decision-making, resulting in improved patient care and reduced costs. Methods: In a prospective clinical trial, patients presenting with acute dyspnea to the ED of the Erasmus Medical College, Rotterdam, the Netherlands, were randomized for either rapid measurement or no measurement of NT-proBNP. For ruling out heart failure, cutoff values of 93 pg/mL in male and 144 pg/mL in female patients were used, and for ruling in heart failure, a cutoff value of 1,017 pg/mL was used. Time to discharge from the hospital and costs related to hospital admission were primary end points. Bootstrap analysis was used for comparison of costs and 30-day mortality between the NT-proBNP and control group. Results: A total of 477 patients (54% male) was enrolled. The mean age was 59 years, with 44% of patients having a history of cardiac disease. Median time to discharge from the hospital was 1.9 days (interquartile range [IQR], 0.12-8.4 days) in the NT-proBNP group (n = 241) compared with 3.9 days (IQR, 0.16-11.0 days) in the control group (n = 236) (P = .04). Introduction of NT-proBNP testing resulted in a trend toward reduction in costs related to hospital admission and diagnostic investigations of $1,364 per patient (95% CI $-246 to $3,215), whereas 30-day mortality was similar (15 patients in the NT-proBNP and 18 patients in the control group). Conclusions: Introduction of NT-proBNP testing for heart failure in the ED setting reduces the time to discharge and is associated with a trend toward cost reduction.

Preconception folic acid treatment affects the microenvironment of the maturing oocyte in humans (Article)

2008-06-01T00:00:00Z

Objective: To investigate the influence of folic acid supplementation on the follicular fluid concentrations of folate and total homocysteine and their relationship to the diameter of the follicle. Design: Observational study. Setting: Tertiary referral fertility clinic at the Erasmus MC, University Medical Center, Rotterdam, The Netherlands. Patient(s): Thirty-seven women undergoing IVF or intracytoplasmic sperm injection treatment. Intervention(s): No interventions other than routine stimulation treatment and the recommendation of folic acid supplementation. Main Outcome Measure(s): Concentrations of folate and total homocysteine in monofollicular and pooled follicular fluid and the diameter of the follicle. Result(s): Folic acid supplementation significantly increased folate and decreased total homocysteine concentrations in pooled follicular fluid. In monofollicular fluid, folate concentrations only were significantly increased in supplemented women. The total homocysteine concentration appeared to be significantly correlated with the diameter of the follicle (r = 0.27). Samples from single follicles were less prone to artifacts in the measurements of the folate and total homocysteine concentration. Conclusion(s): Preconception folic acid supplementation significantly alters both folate and total homocysteine concentrations in follicular fluid. The correlation between the diameter of the follicle and total homocysteine concentration in follicular fluid warrants further investigation.

Eight-fold increased risk for congenital heart defects in children carrying the nicotinamide N-methyltransferase polymorphism and exposed to medicines and low nicotinamide (Article)

2008-06-01T00:00:00Z

Aims: Congenital heart defects (CHDs) have a multifactorial origin, in which subtle genetic factors and peri-conception exposures interact. We hypothesize that derangements in the homocysteine and detoxification pathways, due to a polymorphism in the nicotinamide N-methyltransferase (NNMT) gene, low maternal dietary nicotinamide intake, and medicine use in the peri-conception period, affect CHD risk. Methods and results: In 292 case and 316 control families, maternal peri-conception medicine use and low dietary intake of nicotinamide (≤13.8 mg/day) were independently associated with CHD risk [odds ratio (95% confidence interval) 1.6 (1.1-2.3) and 1.5 (1.03-2.3), respectively]. No significant association was found for the NNMT AG/AA genotype in mothers [0.9 (0.7-1.3)], fathers [1.1 (0.8-1.6)], or children [1.1 (0.8-1.6)]. However, the combination of peri-conception medicine use, low dietary nicotinamide intake, and the NNMT AG/AA genotype in mothers or children showed risk of 2.7 (1.02-8.1) and 8.8 (2.4-32.5), respectively. Conclusion: Children carrying the NNMT A allele face additional CHD risk in combination with peri-conception exposure to medicines and/or a low dietary nicotinamide intake. These findings provide a first set of data against which future studies with larger sample sizes can be compared with.

Polymorphisms in genes related to folate and cobalamin metabolism and the associations with complex birth defects (Article)

2008-06-01T00:00:00Z

Objective: To investigate the associations between biomarkers and genetic variants involved in homocysteine metabolism and the risk of complex birth defects. Methods: Total homocysteine (tHcy), folate, cobalamin, apo-transcobalamin (apo-TC) and apo-haptocorrin (apo-HC) were measured in the amniotic fluid of 82 women who were pregnant with a child having a complex birth defect, such as neural tube defect, cleft lip and/or palate, heart defect or omphalocele, and in 110 women pregnant with a non-malformed child. The determined genotypes of the child comprised of 5, 10-methylenetetrahydrofolate reductase (MTHFR 677C>T, 1298A>C), methionine synthase (MTR 2756A>G), methionine synthase reductase (MTRR 66A>G) and transcobalamin (TCN2 776C>G). Univariate and multivariate logistic regression analyses were performed. Results: Significantly lower cobalamin and higher apo-TC, apo-HC, tHcy and folate concentrations were determined in amniotic fluids of cases compared with controls (p ≤ 0.001). Logistic regression analysis revealed that after adjustment for maternal age, children carrying the MTHFR 677T allele showed a four-fold increased risk of having a complex birth defect, OR (95% CI) = 4.0 (1.1-15.4). Other genotypes did not show significant associations. Conclusion: The MTHFR 677C>T polymorphism in conjunction with reduced folate- and/or cobalamin status may increase the risk of complex birth defects. Copyright

Association of graded allele-specific changes in CYP2D6 function with imipramine dose requirement in a large group of depressed patients (Article)

2008-06-01T00:00:00Z

The inactivation and clearance of the tricyclic antidepressant imipramine is dependent on CYP2D6 activity. First, CYP2C19 converts imipramine into the active metabolite desipramine, which is then inactivated by CYP2D6. This retrospective single center study aimed to prove whether CYP2C19 and ample CYP2D6 genotyping (taking into consideration four null alleles and three decreased-activity alleles) could be used to predict imipramine and desipramine plasma concentrations in depressed patients, and whether genotype-based drug dose recommendations might assist in the early management of imipramine pharmacotherapy. In 181 subjects with major depressive disorder, drug doses were recorded, imipramine and desipramine plasma concentrations were monitored and CYP2C19 (*2) and CYP2D6 genotype (*3, *4, *5, *6, *9, *10, *41 and gene duplication) were obtained, yielding graded allele-specific CYP2D6 patient groups. Desipramine and imipramine+desipramine plasma concentration per drug dose unit, imipramine dose at steady state, and imipramine dose requirement significantly depended on CYP2D6 genotype (Kruskal-Wallis test, P<0.0001). Mean (±s.d.) drug dose requirements were 131 (±109), 155 (±70), 217 (±95), 245 (±125), 326 (±213), and 509 (±292) mg imipramine/day in carriers of 0, 0.5, 1, 1.5, 2, and >2 active CYP2D6 genes, respectively. Our protocol for CYP2D6 genotyping will thus importantly aid in the prediction of imipramine metabolism, allowing for the use of an adjusted starting dose and faster achievement of predefined imipramine+desipramine plasma levels in all genetic patient subgroups. Therefore, therapeutic efficacy and efficiency may be improved, the number of adverse drug reactions decreased, and hospital stay reduced.

Genetic and lifestyle factors related to the periconception vitamin B12 status and congenital heart defects: A Dutch case-control study (Article)

2008-05-01T00:00:00Z

Maternal hyperhomocysteinemia is associated with congenital heart defects (CHDs) in the offspring. A low periconception vitamin B12 status is determined by genetic and lifestyle factors and causes hyperhomocysteinemia. We investigated methionine synthase reductase (MTRR) and transcobalamin II (TC) genes and maternal intake and serum concentrations of vitamin B12 in association with CHD risk. Seventeen months after the index-pregnancy, we studied 230 children with a CHD and 251 non-malformed children and their parents. Data were collected on current and periconception maternal vitamin supplement use and maternal dietary vitamin B12 intake of the month before the study moment. Blood samples were taken for the determination of MTRR A66G and TC C776G genotypes in families and maternal serum vitamin B12 concentrations. Transmission disequilibrium tests and univariate and multivariate analyses were applied. Allele transmissions were not significantly distorted. The MTRR and TC genotypes did not significantly affect CHD risk. Neither polymorphisms in mothers and/or children revealed significant interactions nor in combination with low vitamin B12 intake. Low maternal serum vitamin B12 combined with the maternal or child's MTRR 66 GG genotype resulted in odds ratios of 1.4 (95% confidence interval 0.6-3.5) and 1.3 (0.5-3.4), respectively. The TC 776 GG genotype in mothers and children revealed risk estimates of 2.2 (0.7-7.1) and 1.9 (0.5-7.4), respectively. In conclusion, MTRR 66 GG and TC 776 GG genotypes in mothers and children may contribute to the risk of CHDs, particularly when the maternal vitamin B12 status is low. The future enlargement of our sample size might demonstrate significant associations.

Early Detection of Left Ventricular Dysfunction by Doppler Tissue Imaging and N-terminal Pro-B-type Natriuretic Peptide in Patients with Symptomatic Severe Aortic Stenosis (Article)

2008-03-01T00:00:00Z

Background: Patients with severe aortic stenosis (AS) require valve replacement before development of irreversible left ventricular (LV) dysfunction. It has been postulated that Doppler tissue imaging (DTI) parameters are more sensitive to detect subtle LV dysfunction compared with conventional echocardiographic parameters. Objective: We sought to assess early LV dysfunction with DTI-derived echocardiographic parameters and N-terminal pro-B-type natriuretic peptide (NT-proBNP) in patients with severe AS and normal LV ejection fraction. Methods: A total of 29 patients (mean age 65 ± 12 years, 15 male) with symptomatic severe AS and 17 control subjects were included in the study. DTI was performed at the level of the mitral lateral (mlat) and septal (msep) annulus. Systolic (Sm), early (Em), and late (Am) diastolic velocities were measured, and E/Em ratio was calculated. NT-proBNP was determined by an electrochemiluminescence immunoassay. Results: Baseline characteristics between patients and control subjects were similar regarding LV ejection fraction and mitral inflow E/A ratio. However, patients with AS had significantly lower DTI values (Sm, Em, Am) compared with control subjects. Moreover, LV filling pressures, expressed by the E/Em ratio, were significantly higher in patients. Correlation analysis showed a relationship between the natural logarithm of NT-proBNP and aortic valve area, Smlat, and E/Emsepratio. Using stepwise multiple linear regression, Smlatwas found to be independently related to NT-proBNP. Conclusions: In patients with severe AS and normal LV ejection fraction, DTI showed LV systolic and diastolic dysfunction compared with control subjects. DTI-derived variables, and especially Smlat, were correlated with NT-proBNP levels.

Low folate in seminal plasma is associated with increased sperm DNA damage (Article)

2008-01-01T00:00:00Z

Objective: To determine associations between vitamin B status, homocysteine (tHcy), semen parameters, and sperm DNA damage. Design: Observational study. Setting: A tertiary referral fertility clinic. Patient(s): Two hundred fifty-one men of couples undergoing in vitro fertilization (IVF) or intracytoplasmic sperm injection (ICSI) treatment, with subgroups of fertile (n = 70) and subfertile men (n = 63) defined according to semen concentration and proven fertility. Intervention(s): None. Main Outcome Measure(s): The DNA fragmentation index (DFI) as marker of sperm DNA damage determined using the sperm chromatin structure assay (SCSA), and semen parameters assessed according to World Health Organization criteria; tHcy, folate, cobalamin, and pyridoxine concentrations determined in seminal plasma and blood. Result(s): In the total group of fertile and subfertile men, all biomarkers in blood were statistically significantly correlated with those in seminal plasma. No correlation was found between the biomarkers in blood and the semen parameters. In seminal plasma, both tHcy and cobalamin positively correlated with sperm count. Folate, cobalamin, and pyridoxine were inversely correlated with ejaculate volume. In fertile men, seminal plasma folate showed an inverse correlation with the DNA fragmentation index. Conclusion(s): Low concentrations of folate in seminal plasma may be detrimental for sperm DNA stability.

Influence of Renal Function on the Usefulness of N-Terminal Pro-B-Type Natriuretic Peptide as a Prognostic Cardiac Risk Marker in Patients Undergoing Noncardiac Vascular Surgery (Article)

2008-01-01T00:00:00Z

N-terminal pro-B-type natriuretic peptide (NT-pro-BNP) is related to stress-induced myocardial ischemia and/or volume overload, both common in patients with renal dysfunction. This might compromise the prognostic usefulness of NT-pro-BNP in patients with renal impairment before vascular surgery. We assessed the prognostic value of NT-pro-BNP in the entire strata of renal function. In 356 patients (median age 69 years, 77% men), cardiac history, glomerular filtration rate (GFR, ml/min/1.73 m2), and NT-pro-BNP level (pg/ml) were assessed preoperatively. Troponin T and electrocardiography were assessed postoperatively on days 1, 3, 7, and 30. The end point was the composite of cardiovascular death, Q-wave myocardial infarction, and troponin T release. Multivariate analysis was used to evaluate the interaction between GFR, NT-pro-BNP and their association with postoperative outcome. Median GFR was 78 ml/min/1.73 m2and the median concentration of NT-pro-BNP was 197 pg/ml. The end point was reached in 64 patients (18%); cardiac death occurred in 7 (2.0%), Q-wave myocardial infarction in 34 (9.6%), and non-Q-wave myocardial infarction in 23 (6.5%). After adjustment for confounders, NT-pro-BNP levels and GFR remained significantly associated with the end point (p = 0.005). The prognostic value of NT-pro-BNP was most pronounced in patients with GFR ≥90 (odds ratio [OR] 1.18, 95% confidence interval [CI] 0.80 to 1.76) compared with patients with GFR 60 to 89 (OR 1.04, 95% CI 1.002 to 1.07), and with GFR 30 to 59 (OR 1.12, 95% CI 1.03 to 1.21). In patients with GFR <30 ml/min/1.73 m2, NT-pro-BNP levels have no prognostic value (OR 1.00, 95% CI 0.99 to 1.01). In conclusion, the discriminative value of NT-pro-BNP is most pronounced in patients with GFR ≥90 ml/min/1.73 m2and has no prognostic value in patients with GFR <30 ml/min/1.73 m2.

Low dietary riboflavin but not folate predicts increased fracture risk in postmenopausal women homozygous for the MTHFR 677 T allele (Article)

2008-01-01T00:00:00Z

The MTHFR C677T polymorphism is associated with mildly elevated homocysteine levels when folate and/or riboflavin status is low. Furthermore, a mildly elevated homocysteine level is a risk factor for osteoporotic fractures. We studied whether dietary intake of riboflavin and folate modifies the effects of the MTHFR C677T variant on fracture risk in 5035 men and women from the Rotterdam Study. We found that the MTHFR C677T variant interacts with dietary riboflavin intake to influence fracture risk in women. Introduction: The MTHFR C677T polymorphism is associated with mildly elevated homocysteine (Hcy) levels in the presence of low folate and/or riboflavin status. A mildly elevated Hcy level was recently identified as a modifiable risk factor for osteoporotic fracture. We studied whether dietary intake of riboflavin and folate modifies the effects of the MTHFR C677T polymorphism on BMD and fracture risk. Materials and Methods: We studied 5035 individuals from the Rotterdam Study, ≥55 yr of age, who had data available on MTHFR, nutrient intake, and fracture risk. We performed analysis on Hcy levels in a total of 666 individuals, whereas BMD data were present for 4646 individuals (2692women). Results: In the total population, neither the MTHFR C677T polymorphism nor low riboflavin intake was associated with fracture risk and BMD. However, in the lowest quartile of riboflavin intake, female 677-T homozygotes had a 1.8 (95% CI: 1.1-2.9, p = 0.01) times higher risk for incident osteoporotic fractures and a 2.6 (95% CI: 1.3-5.1, p = 0.01) times higher risk for fragility fractures compared with the 677-CC genotype (interaction, p = 0.0002). This effect was not seen for baseline BMD in both men and women. No significant influence was found for dietary folate intake on the association between the MTHFR C677T genotype and fracture risk or BMD. In the lowest quartile of dietary riboflavin intake, T-homozygous individuals (men and women combined) had higher (22.5%) Hcy levels compared with C-homozygotes (mean difference = 3.44 μM, p = 0. 01; trend, p = 0.02). Conclusions: In this cohort of elderly whites, the MTHFR C677T variant interacts with dietary riboflavin intake to influence fracture risk in women.

The Generation R Study Biobank: A resource for epidemiological studies in children and their parents (Article)

2007-12-01T00:00:00Z

The Generation R Study is a population-based prospective cohort study from fetal life until young adulthood. The study is designed to identify early environmental and genetic causes of normal and abnormal growth, development and health from fetal life until young adulthood. In total, 9,778 mothers were enrolled in the study. Prenatal and postnatal data collection is conducted by physical examinations, questionnaires, interviews, ultrasound examinations and biological samples. Major efforts have been conducted for collecting biological specimens including DNA, blood for phenotypes and urine samples. In this paper, the collection, processing and storage of these biological specimens are described. Together with detailed phenotype measurements, these biological specimens form a unique resource for epidemiological studies focused on environmental exposures, genetic determinants and their interactions in relation to growth, health and development from fetal life onwards.

Hypokalaemia and subsequent hyperkalaemia in hospitalized patients (Article)

2007-12-01T00:00:00Z

Background. The objective was to study the epidemiology of hypokalaemia [serum potassium concentration (SK) <3.5 mmol/l] in a general hospital population, specifically focusing on how often and why patients develop subsequent hyperkalaemia (SK<5.0 mmol/l). Methods. In a 3-month hospital-wide study we analysed factors contributing to hypokalaemia and subsequent hyperkalaemia. Results. From 1178 patients in whom SKwas measured, 140 patients (12%) with hypokalaemia were identified (SK3.0 ± 0.3 mmol/l). One hundred patients (71%) had hospital-acquired hypokalaemia. Common causes of hypokalaemia included gastrointestinal losses (67%), diuretics (36%) and haematological malignancies (9%). In 104 patients (74%), hypokalaemia was multifactorial. Hypokalaemia frequently coexisted with hyponatraemia (24%) and, when measured, hypomagnesaemia (61%). Twenty-three patients (16%) developed hyperkalaemia (highest SK5.7 ± 0.7 mmol/l) following hypokalaemia. In these patients, potassium suppletion was not more common (70 vs 59%, P = 0.5), but when potassium was given, the total amount administered was significantly higher (median 350 mmol vs 180 mmol, P = 0.02). Furthermore, these patients more often received total parenteral nutrition (17 vs 4%, P = 0.02) and magnesium suppletion (30 vs 9%, P = 0.009), and more often had haematological malignancies (22 vs 6%, P = 0.03). Conclusions. Hypokalaemia is a multifactorial and usually hospital-acquired condition associated with hyponatraemia and hypomagnesaemia. One out of every six patients with hypokalaemia developed subsequent hyperkalaemia. Besides potassium suppletion, total parenteral nutrition (source of potassium), magnesium suppletion (may reduce kaliuresis) and haematological malignancy (may cause cell lysis) contribute to hyperkalaemia following hypokalaemia. Caution with potassium suppletion and frequent monitoring of SKmay prevent iatrogenic hyperkalaemia.

Baseline natriuretic peptide levels in relation to myocardial ischemia, troponin T release and heart rate variability in patients undergoing major vascular surgery (Article)

2007-12-01T00:00:00Z

BACKGROUND: This study was conducted to determine the association between baseline N-terminal pro-B-type natriuretic peptide (NT-proBNP) and myocardial ischemia, troponin T release and heart rate variability (HRV) in patients undergoing major vascular surgery. METHODS: In a prospective study, 182 vascular surgery patients were evaluated by clinical risk factors, dobutamine stress echocardiography and baseline NT-proBNP levels. Myocardial ischemia was detected by continuous 12-lead electrocardiographic monitoring starting 1 day before to 2 days after surgery. Troponin T (>0.03 ng/ml) was measured on day 1, 3 and 7 postoperatively and before discharge. HRV was measured at the day prior to surgery. RESULTS: The median NT-proBNP level was 184 ng/l (interquartile range: 79-483 ng/l). Myocardial ischemia was detected in 21% and troponin T release in 17% of patients. After adjustment for clinical risk factors and stress echocardiography results, higher NT-proBNP levels (per 1 ng/l increase in the natural logarithm of NT-proBNP) were associated with a higher incidence of myocardial ischemia (odds ratio: 1.59, 95% confidence interval: 1.21-2.08, P<0.001) and troponin T release (odds ratio: 1.76, 95% confidence interval: 1.33-2.34, P<0.001). The optimal cutoff value of NT-proBNP to predict ischemia and/or troponin T release was 270 ng/l (area under the curve: 0.70). Higher baseline NT-proBNP levels were also associated with a larger ischemic burden at electrocardiographic monitoring (r=0.22, P=0.03). No significant correlation, however, was found between NT-proBNP and preoperative HRV (r=-0.024, P=0.78). CONCLUSION: Elevated baseline NT-proBNP levels are significantly associated with perioperative myocardial ischemia and troponin T release, but not with preoperative HRV in patients undergoing major vascular surgery.

Seminal plasma cobalamin significantly correlates with sperm concentration in men undergoing IVF or ICSI procedures (Article)

2007-07-01T00:00:00Z

Mild hyperhomocysteinemia is caused by B vitamin deficiencies. We hypothesize that these biochemical derangements detrimentally affect spermatogenesis. Therefore, the aim of this study was to investigate the folate, cobalamin, pyridoxine, and homocysteine concentrations in blood and seminal plasma and the associations between these biomarkers and semen parameters in men participating in an in vitro fertilization or intracytoplasmic sperm injection program. From 73 men (median age [range]: 37 years [28-53]), blood and semen samples were obtained for the determination of serum and red blood cell (RBC) folate, serum total cobalamin, whole-blood pyridoxal-5′-phosphate, plasma total homocysteine (tHcy), and serum total testosterone. Semen analysis included sperm concentration, motility, and morphology according to World Health Organization criteria. The B vitamins and tHcy concentrations were significantly correlated in blood but not in seminal plasma. The serum and RBC folate concentrations were significantly correlated also with the total folate concentration in seminal plasma (r = .44; P < .001 and r = .39; P < .001, respectively). Likewise, the total cobalamin concentration in serum and seminal plasma was significantly correlated (r = .55; P = .001). Of interest is that the total cobalamin concentration in seminal plasma was significantly correlated with the sperm concentration (r= .42; P< .001). This is in contrast to the absence of significant associations between the other vitamins and tHcy in blood and seminal plasma and any of the semen parameters. These findings suggest that folate and cobalamin are transferred from the blood to the male reproductive organs and emphasize the role of cobalamin in spermatogenesis in human. Copyright

Validation of the assessment of folate and vitamin B12 intake in women of reproductive age: The method of triads (Article)

2007-05-01T00:00:00Z

Objective: To validate the folate and vitamin B12intakes estimated by a food-frequency questionnaire (FFQ) designed to be used in a case-control study on the association between maternal dietary intake and the risk of having a child with a congenital heart defect. Design and subjects: The FFQ was filled out by 53 women of reproductive age. Immediately thereafter, blood samples were taken to determine serum folate, red blood cell (RBC) folate and serum vitamin B12concentrations. Subsequently, three dietary 24-h recalls (24HR) were completed during a period of three successive weeks and used as a reference method. The recalls comprised two weekdays and one weekend day. Using the method of triads, validity coefficients were calculated by comparing nutrient intakes derived from the FFQ and 24HR with the corresponding nutritional biomarkers in blood. The validity coefficient is the correlation between the dietary intake reported by the FFQ and the unknown 'true' dietary intake. Results: The comparison of B-vitaminin takes reported by the FFQ and the mean of the 24HR revealed deattenuated correlation coefficients of 0.98 for folate and 0.66 for vitamin B12. The correlation coefficients between the B-vitamin intakes estimated by the FFQ and concentrations of serum folate, RBC folate and serum vitamin B12were 0.20, 0.28 and 0.21, respectively. The validity coefficients for serum folate, RBC folate and serum vitamin B12were 0.94, 0.75 and 1.00, respectively. The estimated folate and vitamin B12intakes were comparable with the results of the most recent Dutch food consumption survey. Conclusions: The adapted FFQ is a reliable tool to estimate the dietary intake of energy, macronutrients, folate and vitamin B12in women of reproductive age. Therefore, this FFQ is suitable for the investigation of nutrient-disease associations in future.

Novel risk factors for hospital-acquired hyponatraemia: A matched case-control study (Article)

2007-03-01T00:00:00Z

Background: Hospital-acquired hyponatraemia is a common and potentially serious condition. Risk factors for hospital-acquired hyponatraemia have not been studied in a controlled fashion. Methods: From 1501 patients in whom serum sodium (SNa) was determined, 50 cases with hospital-acquired hyponatraemia (in-hospital decrease in SNa≥ 7 mmol/l to < 136 mmol/l) were identified. They were matched by age, gender and department to 69 normonatraemic controls. Results: In the 50 cases, SNafell from 141 ± 2 to 130 ± 4 mmol/l, while controls remained normonatraemic. During the development of hyponatraemia, C-reactive protein (CRP) increased in cases (median from 23 to 146 mg/l), whereas it decreased in controls (median from 31 to 24 mg/l, P = 0.008). Additional factors associated with hospital-acquired hyponatraemia included diabetes mellitus (16/50 vs. 10/69, P = 0.009) and the use of insulin (12/50 vs. 4/69, P = 0.007), antibiotics (41/50 vs. 38/69, P = 0.006) and opioids (32/50 vs. 27/69, P = 0.005). Multivariate conditional logistic regression showed that the use of insulin [odds ratio (OR) 10.5, 95% confidence interval (CI) 1.5-72.4], antibiotics (OR 4.5, 95% CI 1.4-14.6) and opioids (OR 2.9, 95% CI 1.1-7.8) was also independently associated with hospital-acquired hyponatraemia. Mortality (6/50 vs. 1/69, P = 0.04) and intensive care admission (15/50 vs. 7/69, P = 0.008) were higher in cases. Conclusions: An increase in CRP and the use of insulin, antibiotics and opioids are novel risk factors for hospital-acquired hyponatraemia. These factors represent interesting new clues regarding the pathophysiology of hospital-acquired hyponatraemia, suggesting that the acute-phase response, pain and/or direct drug effects could be involved in the release of antidiuretic hormone.

Long-term toxicity of [177Lu-DOTA0,Tyr 3]octreotate in rats (Article)

2007-02-01T00:00:00Z

Purpose and methods: Studies on peptide receptor radionuclide therapy (PRRT) using radiolabelled somatostatin analogues have shown promising results with regard to tumour control. The efficacy of PRRT is limited by uptake and retention in the proximal tubules of the kidney, which might lead to radiation nephropathy. We investigated the long-term renal toxicity after different doses of [177Lu-DOTA0,Tyr3]octreotate and the effects of dose fractionation and lysine co-injection in two tumour-bearing rat models. Results: Significant renal toxicity was detected beyond 100 days after start of treatment as shown by elevated serum creatinine and proteinuria. Microscopically, tubules were strongly dilated with flat epithelium, containing protein cylinders. Creatinine levels rose significantly after 555 MBq [177Lu-DOTA0,Tyr3]octreotate, but were significantly lower after 278 MBq (single injection) or two weekly doses of 278 MBq. Renal damage scores were maximal after 555 MBq and significantly lower in the 278 and 2x278 MBq groups. Three doses of 185 MBq [177Lu-DOTA0,Tyr3]octreotate with intervals of a day, a week or a month significantly influenced serum creatinine (469±18, 134±70 and 65±15 μmol/l, respectively; p<0.001). Renal histological damage scores were not significantly influenced by dose fractionation. Lysine co-administration with three weekly treatments of 185 MBq significantly lowered serum creatinine and proteinuria. Conclusion: Injection of high doses of [177Lu-DOTA0,Tyr3]octreotate resulted in severe renal damage in rats as indicated by proteinuria, elevated serum creatinine and histological damage. This damage was dose dependent and became overt between 100 and 200 days after treatment. Dose fractionation had significant beneficial effects on kidney function. Also, lysine co-injection successfully prevented functional damage.

Klinische chemie: van labstraat naar zorgpad (Inaugural Lecture)

2005-09-09T00:00:00Z

A common polymorphism in the CYP3A7 gene is associated with a nearly 50% reduction in serum dehydroepiandrosterone sulfate levels. (Article)

2005-09-01T00:00:00Z

CONTEXT: CYP3A7, expressed in the human fetal liver and normally silenced after birth, plays a major role in the 16alpha-hydroxylation of dehydroepiandrosterone (DHEA), DHEA sulfate (DHEAS), and estrone. Due to a replacement of part of the CYP3A7 promoter with a sequence identical with the same region in the CYP3A4 promoter (referred to as CYP3A7*1C), some individuals still express a variant of the CYP3A7 gene later in life. OBJECTIVE: The objective of this study was to examine the effect of the CYP3A7*1C polymorphism on serum steroid hormone levels. DESIGN, SETTING, PARTICIPANTS: Two population-based cohort studies were performed. Study group 1 consisted of 208 subjects randomly selected from the Rotterdam Study, and study group 2 consisted of 345 elderly independently living men. MAIN OUTCOME MEASURES: Serum DHEA(S), androstenedione, estradiol, estrone, and testosterone levels were the main outcome measures. RESULTS: In study groups 1 and 2, heterozygous CYP3A7*1C carriers had almost 50% lower DHEAS levels compared with homozygous carriers of the reference allele [study group 1, 1.74 +/- 0.25 vs. 3.33 +/- 0.15 micromol/liter (P = 0.02); study group 2, 2.09 +/- 0.08 vs. 1.08 +/- 0.12 micromol/liter (P < 0.001)]. No differences in circulating DHEA, androstenedione, estradiol, or testosterone levels were found. However, in study group 2, serum estrone levels were lower in heterozygous CYP3A7*1C carriers compared with homozygous carriers of the reference allele (0.11 +/- 0.002 vs. 0.08 +/- 0.006 nmol/liter; P < 0.001). CONCLUSION: The CYP3A7*1C polymorphism causes the persistence of enzymatic activity of CYP3A7 during adult life, resulting in lower circulating DHEAS and estrone levels.

A shift to a peripheral Th2-type cytokine pattern during the carcinogenesis of cervical cancer becomes manifest in CIN III lesions (Article)

2005-01-01T00:00:00Z

BACKGROUND: A shifted balance between T helper 1 (Th1)-type and Th2-type cytokines has been hypothesised in cervical dysplasia. AIMS: To evaluate possible deregulation of the cytokine network by estimating the expression of peripheral cytokines in different stages of cervical disease and in relation to the presence or absence of high risk human papillomavirus (HR-HPV). METHODS: Twenty one HR-HPV positive women with high grade cervical intraepithelial neoplasia (CIN II-III) and 12 patients with invasive cervical carcinoma formed the study groups. Two control groups consisted of 10 HR-HPV positive and 11 HR-HPV negative women without CIN. Differences in leucocyte subgroups were evaluated by a differential leucocyte count. Plasma concentrations of tumour necrosis factor alpha (TNFalpha), TNFalpha receptors TNFRI and TNFRII, interferon gamma (IFNgamma), interleukin 2 (IL-2), IL-12, IL-4, and IL-10 were determined by enzyme linked immunosorbent assays. RESULTS: Leucocyte counts in patients with CIN III and carcinoma were significantly higher than in controls. Plasma IFNgamma concentrations were significantly lower in patients with CIN III and carcinoma than in women with CIN II or controls. Plasma concentrations of IL-12, IL-2, IL-4, and TNFalpha did not differ significantly between groups, but significantly lower plasma concentrations of TNFRII were found in CIN III and carcinoma compared with CIN II. IL-10 was detected with increased frequency in the plasma of patients with CIN III and carcinoma. CONCLUSIONS: These results indicate that a shift to a Th2-type cytokine pattern during the carcinogenesis of cervical cancer occurs in women with CIN III lesions.

Automated counting of white blood cells in synovial fluid. (Article)

2004-02-01T00:00:00Z

OBJECTIVES: To evaluate the performance of automated leucocyte (white blood cell; WBC) counting by comparison with manual counting. METHODS: The number of WBC was determined in heparinized synovial fluid samples by the use of (i) a standard urine cytometer (Kova) and a microscope (reference method) and (ii) a haematology analyser (Sysmex XE-2100; WBC/BASO and DIFF channels). Imprecision within and between days was determined by replicate analysis of a low (level A; WBC approximately 0.560 x 10(9)/l) and a high (level B; WBC approximately 1.081 x 10(9)/l) dedicated synovial fluid control (Quantimetrix). RESULTS: The WBC count of the DIFF channel was highly correlated with the WBC count of the microscopic reference method (r = 0.99; WBC analyser = 0.870 x WBC reference method + 0.413). In contrast, no agreement existed between WBC counts generated by the WBC/BASO channel of the analyser and the reference method (r = 0.52; WBC analyser = 0.008 x WBC reference method + 0.079). Within-day imprecision (4-7%) and between-day imprecision (10%) of the haematology analyser were smaller than the within-day imprecision (12%) and the between-day imprecision (20-22%) of the manual reference method. For manual counting, inter-observer coefficients of variation were 35.9% (control level A) and 21.0% (control level B). CONCLUSIONS: The WBC count in synovial fluid can be reliably determined using the DIFF channel of the Sysmex XE-2100. Automated counting of WBC in synovial fluid offers more precise and faster results than manual counting.

Sequencing: not always the "gold standard". (Article)

2004-01-01T00:00:00Z

Homocysteine levels and the risk of osteoporotic fracture (Article)

2004-01-01T00:00:00Z

BACKGROUND: Very high plasma homocysteine levels are characteristic of homocystinuria, a rare autosomal recessive disease accompanied by the early onset of generalized osteoporosis. We therefore hypothesized that mildly elevated homocysteine levels might be related to age-related osteoporotic fractures. METHODS: We studied the association between circulating homocysteine levels and the risk of incident osteoporotic fracture in 2406 subjects, 55 years of age or older, who participated in two separate prospective, population-based studies. In the Rotterdam Study, there were two independent cohorts: 562 subjects in cohort 1, with a mean follow-up period of 8.1 years; and 553 subjects in cohort 2, with a mean follow-up period of 5.7 years. In the Longitudinal Aging Study Amsterdam, there was a single cohort of 1291 subjects, with a mean follow-up period of 2.7 years. Multivariate Cox proportional-hazards regression models were used for analysis of the risk of fracture, with adjustment for age, sex, body-mass index, and other characteristics that may be associated with the risk of fracture or with increased homocysteine levels. RESULTS: During 11,253 person-years of follow-up, osteoporotic fractures occurred in 191 subjects. The overall multivariable-adjusted relative risk of fracture was 1.4 (95 percent confidence interval, 1.2 to 1.6) for each increase of 1 SD in the natural-log-transformed homocysteine level. The risk was similar in all three cohorts studied, and it was also similar in men and women. A homocysteine level in the highest age-specific quartile was associated with an increase by a factor of 1.9 in the risk of fracture (95 percent confidence interval, 1.4 to 2.6). The associations between homocysteine levels and the risk of fracture appeared to be independent of bone mineral density and other potential risk factors for fracture. CONCLUSIONS: An increased homocysteine level appears to be a strong and independent risk factor for osteoporotic fractures in older men and women.

CYP3A5 variant allele frequencies in Dutch Caucasians (Article)

2002-01-01T00:00:00Z

BACKGROUND: Enzymes of the cytochrome P450 3A (CYP3A) family are responsible for the metabolism of >50% of currently prescribed drugs. CYP3A5 is expressed in a limited number of individuals. The absence of CYP3A5 expression in approximately 70% of Caucasians was recently correlated to a genetic polymorphism (CYP3A5*3). Because CYP3A5 may represent up to 50% of total CYP3A protein in individuals polymorphically expressing CYP3A5, it may have a major role in variation of CYP3A-mediated drug metabolism. Using sequencing, have been identified (Hustert et al. Pharmacogenetics 2001;11:773-9; Kuehl et al. Nat Genet 2001;27:383-91) variant alleles *2 through *7 for CYP3A5. Detection of CYP3A5 variant alleles, and knowledge about their allelic frequency in specific ethnic groups, is important to establish the clinical relevance of screening for these polymorphisms to optimize pharmacotherapy. METHODS: In a group of 500 healthy Dutch Caucasian blood donors, we determined the allelic frequency of the CYP3A5*2, *3, *4, *5, *6, and *7 alleles by use of newly developed PCR-restriction fragment length polymorphism assays. RESULTS: The frequency of the defective CYP3A5*3 allele in the Dutch Caucasian population was 91%, followed by the CYP3A5*2 (1%) and CYP3A5*6 (0.1%) alleles. The CYP3A5*4, *5, and *7 alleles were not detected. CONCLUSIONS: On the basis of its allelic frequency, screening for the CYP3A5*3 allele in the Caucasian population is extremely relevant. In addition, screening for the CYP3A5*2 allele may be taken into consideration in individuals heterozygous for the CYP3A5*3 allele. The CYP3A5*4, *5, *6, and *7 alleles have low allelic frequencies that do not support initial screening.

The CYP3A4*3 allele: is it really rare? (Article)

2001-01-01T00:00:00Z

CYP3A4-V polymorphism detection by PCR-restriction fragment length polymorphism analysis and its allelic frequency among 199 Dutch Caucasians (Article)

2000-01-01T00:00:00Z

Homocysteine and short-term risk of myocardial infarction and stroke in the elderly: the Rotterdam Study (Article)

1999-01-01T00:00:00Z

BACKGROUND: Elevated homocysteine level increases vascular disease risk. Most data are based on subjects younger than 60 years; data for the elderly are more limited. We examined the relationship of homocysteine level to incident myocardial infarction and stroke among older subjects in a nested case-control study. METHODS: Subjects were participants in the Rotterdam Study, a cohort study among 7983 subjects residing in the Ommoord district of Rotterdam, the Netherlands. Baseline examinations were performed from March 1, 1990, to July 31, 1993. The analysis is restricted to myocardial infarction and stroke that occurred before December 31, 1994. One hundred four patients with a myocardial infarction and 120 with a stroke were identified with complete data. Control subjects consisted of a sample of 533 subjects drawn from the study base, free of myocardial infarction and stroke. Nonfasting total homocysteine levels were measured. RESULTS: Results were adjusted for age and sex. The risk of stroke and myocardial infarction increased directly with total homocysteine. The linear coefficient suggested a risk increase by 6% to 7% for every 1-micromol/L increase in total homocysteine. The risk by quintiles of total homocysteine level was significantly increased only in the group with levels above 18.6 micromol/L (upper quintile): odds ratios were 2.43 (95% confidence interval, 1.11-5.35) for myocardial infarction and 2.53 (95% confidence interval, 1.19-5.35) for stroke. Associations were more pronounced among those with hypertension. CONCLUSIONS: The present study, based on a relatively short follow-up period, provides evidence that among elderly subjects an elevated homocysteine level is associated with an increased risk of cardiovascular disease.

Portable blood gas and electrolyte analyzer evaluated in a multiinstitutional study (Article)

1999-01-01T00:00:00Z

A recently introduced blood gas/electrolyte analyzer (SenDx 100((R)), renamed ABL70) intended for point-of-care, near-patient, or stat laboratory use was evaluated simultaneously in four different institutions and compared with three different laboratory bench analyzers with respect to imprecision, inaccuracy (assessed by tonometry), and patient-sample analyses. The analyzer is equipped with a sensor cassette and a reagent cartridge for 50, 100, or 200 analyses and 100 or more traditional quality-control measurements. One analysis requires 170 microL of whole blood and takes <90 s. Statistically, the instrument performed somewhat better (lower CVs) for PO2 and potassium and somewhat worse for pH, PCO2, and ionized calcium than the respective comparison analyzers. However, the overall performance (in terms of CV and accuracy) was satisfactory in terms of clinical (e.g., CLIA '88) goals in all institutions. The mean difference and the CV of that difference in some 400 patient-sample comparisons were as follows: 0.010 (+/- 0.002%) for pH, -0.65 mmHg (+/- 4%) for PCO2, -0.49 mmHg (+/- 6%) for Po2, 0.44 mmol/L (+/- 1.2%) for sodium, -0.013 mmol/L (+/- 2.9%) for potassium, -0.016 mmol/L (+/- 2.6%) for ionized calcium, and -0.016 L/L (+/- 7. 1%) for the hematocrit. Its acceptable analytical performance and ease of operation make the SenDx 100 suitable for the analysis of blood gases and electrolytes.

Optimization of apolipoprotein(a) genotyping with pulsed field gel electrophoresis (Article)

1999-01-01T00:00:00Z

BACKGROUND: Increased lipoprotein(a) is a risk factor for atherosclerosis, and its concentration in serum is inversely correlated with the size of the apoliprotein(a) [apo(a)] component. The size of the apo(a) gene is determined mainly by the Kringle IV size polymorphism. We have optimized and characterized pulsed field gel electrophoresis (PFGE) for apo(a) genotyping. METHODS: Established PFGE protocols were adjusted. The changes included the following: (a) increased DNA yields by the use of all leukocytes for isolation from either 3 mL of fresh EDTA whole blood or 250 microL of frozen buffy coats; (b) increased efficiency of Kpn1 digestion by the inclusion of a digestion buffer wash; (c) reduction of assay time by the use of capillary blotting; (d) increased sensitivity by the use of four digoxigenin-labeled apo(a) probes; and (e) identification using a single film by the inclusion of a digoxigenin-labeled lambda marker probe in addition to apo(a) probes in the hybridization mix. RESULTS: In older Caucasians, 93% (buffy coats, n=468) were heterozygous for apo(a) gene size. An inverse correlation between serum lipoprotein(a) and the sum of Kringle IV alleles was found (y = -23x + 1553; r = -0.442; n = 468). Gel-to-gel variation was minimal (3%). Imprecision (SD) was one Kringle IV repeat (control sample containing eight fragments of 72-233 kb; n=34 electrophoretic runs). CONCLUSIONS: The practicality and sensitivity of the apo(a) genotyping technique by PFGE were improved, and accuracy and reproducibility were preserved. The optimized procedure is promising for apo(a) genotyping on frozen buffy coats from large epidemiological studies.

Serum ferritin and risk of myocardial infarction in the elderly: the Rotterdam Study (Article)

1999-01-01T00:00:00Z

BACKGROUND: Elevated body iron stores have been suggested to be a risk factor for ischemic heart disease. OBJECTIVE: We examined whether elevated serum ferritin concentrations, other indicators of iron status, and dietary iron affected the incidence of myocardial infarction (MI) in an elderly population. DESIGN: A nested, case-control study of 60 patients who had their first MI and 112 age- and sex-matched control subjects embedded in the population-based cohort of the Rotterdam Study. RESULTS: The age- and sex-adjusted risk of MI for subjects with serum ferritin concentrations > or = 200 microg/L was 1.82 (95% CI: 0.90, 3.69; P = 0.096). The odds ratio (OR) was 1.26 (95% CI: 0.98, 1.64; P = 0.078) for the highest tertile of serum ferritin and was only slightly altered in a multivariate model. Risk of MI associated with the highest tertile of ferritin was most evident in current or former smokers (OR: 1.68; 95% CI: 1.17, 2.47; P for trend = 0.008) and in subjects with hypercholesterolemia (OR: 1.43; 95% CI: 0.99, 2.11; P for trend = 0.056) or diabetes (OR: 2.41; 95% CI: 1.12, 7.67; P for trend = 0.027). No association with risk of MI was observed for tertiles of serum iron, serum transferrin, or total dietary iron. For dietary heme iron, risk of MI was significantly increased in a multivariate model in which dietary energy, fat, saturated fat, and cholesterol were adjusted for (OR: 4.01; 95% CI: 1.17, 15.87; P for trend = 0.031). CONCLUSION: In the presence of other risk factors, serum ferritin may adversely affect ischemic heart disease risk in the elderly.

Chromogranin A as serum marker for neuroendocrine neoplasia: comparison with neuron-specific enolase and the alpha-subunit of glycoprotein hormones (Article)

1997-01-01T00:00:00Z

Chromogranin A (CgA) is gaining acceptance as a serum marker of neuroendocrine tumors. Its specificity in differentiating between neuroendocrine and nonneuroendocrine tumors, its sensitivity to detect small tumors, and its clinical value, compared with other neuroendocrine markers, have not clearly been defined, however. The objectives of this study were to evaluate the clinical usefulness of CgA as neuroendocrine serum marker. Serum levels of CgA, neuron-specific enolase (NSE), and the alpha-subunit of glycoprotein hormones (alpha-SU) were determined in 211 patients with neuroendocrine tumors and 180 control subjects with nonendocrine tumors. The concentrations of CgA, NSE, and alpha-SU were elevated in 50%, 43%, and 24% of patients with neuroendocrine tumors, respectively. Serum CgA was most frequently increased in subjects with gastrinomas (100%), pheochromocytomas (89%), carcinoid tumors (80%), nonfunctioning tumors of the endocrine pancreas (69%), and medullary thyroid carcinomas (50%). The highest levels were observed in subjects with carcinoid tumors. NSE was most frequently elevated in patients with small cell lung carcinoma (74%), and alpha-SU was most frequently elevated in patients with carcinoid tumors (39%). Most subjects with elevated alpha-SU levels also had elevated CgA concentrations. A significant positive relationship was demonstrated between the tumor load and serum CgA levels (P < 0.01, by chi 2 test). Elevated concentrations of CgA, NSE, and alpha-SU were present in, respectively, 7%, 35%, and 15% of control subjects. Markedly elevated serum levels of CgA, exceeding 300 micrograms/L, were observed in only 2% of control patients (n = 3) compared to 40% of patients with neuroendocrine tumors (n = 76). We conclude that CgA is the best general neuroendocrine serum marker available. It has the highest specificity for the detection of neuroendocrine tumors compared to the other neuroendocrine markers, NSE and alpha-SU. Elevated levels are strongly correlated with tumor volume; therefore, small tumors may go undetected. Although its specificity cannot compete with that of the specific hormonal secretion products of most neuroendocrine tumors, it can have useful clinical applications in subjects with neuroendocrine tumors for whom either no marker is available or the marker is inconvenient for routine clinical use.

Significance of various parameters derived from biological variability of lipoprotein(a), homocysteine, cysteine, and total antioxidant status (Article)

1997-01-01T00:00:00Z

Analytical and biological components of variability and various derived indices have been determined for lipoprotein(a) [Lp(a)], homocysteine (Hcy), cysteine (Cys), and total antioxidant status (TAOS) in ostensibly healthy adult Caucasians and in stable outpatients with an increased serum Lp(a). In healthy Caucasians, average intraindividual biological CVs (CVb) were 20.0% for Lp(a), 9.4% for Hcy, 5.9% for Cys, and 2.8% for TAOS, CVbs being similar in men and women. In the outpatient group, CVbs were comparable for Hcy, Cys, and TAOS, but significantly lower for Lp(a) (7.5% vs 20.0%; P <0.0001). Moreover, a significant inverse relation between both biological and analytical CVs (CVa) and serum Lp(a) concentrations was demonstrated. We conclude that average CVa and CVb values, and hence average derived indices, are adequate for Hcy, Cys, and TAOS, whereas individual values should be used for Lp(a).

Transcobalamin II-mediated uptake of vitamin B12 by rat liver cells (Doctoral Thesis)

1979-05-23T00:00:00Z

Vitamin B12 plays a unique role in mammalian metabolism not only because, as a coenzyme, it is involved in two completely different and unrelated biochemical pathways, - the synthesis of nucleic acid precursors and the catabolism of some fatty acids -, but even more because it gives an excellent example how different groups of living organisms work together and depend on each other for the supply of vital nutrients. Vitamin B12 is almost exclusively found in animal products. However, it is not synthesized by the animals themselves but, they are able in one or the other way to absorb vitamin B12 which is produced bv microorganisms. For instance in ruminants the bacteria in the rumen are the source of the vitamin, which is taken up by the gut, distributed over the tissues and which is subsequently consumed by man with the meat or with the milk. However, the quantity of vitamin B12 , which is available in the food. is so low, that it would be lost if not an elaborate system of carrier proteins and cellular receotor mechanisms selectively collected it from the food and delivered it to the tissues. Intrinsic factor, produced and secreted by the qastric mucosa, binds the vitamin, which enters the body with the food, and hands it over to the ileal mucosa cells, which carry specific receptors for this protein. When the vitamin enters the blood, the plasma transport proteins, the transcobalamins, take it up immediately and deliver it to the tissues.